The long-term objective is to determine the in vivo function, regulation, and toxicological implications of human microsomal cytochrome P450 (CYP) enzymes. Microsomal CYPs metabolize a wide variety of endogenous as well as xenobiotic compounds, including drugs, environmental chemicals, and dietary components. While it is important to study human CYPs, because of the known species differences between humans and laboratory animals in CYP function and regulation, the in vivo function and regulation of human CYPs are difficult to study because of limits on human experimentation, and the lack of suitable cell-based models for most organ systems, particularly for extrahepatic tissues. Thus, our focus has been on developing novel transgenic and knockout mouse models for studying the in vivo function and regulation of human CYPs. Transgenic mice expressing human CYP genes (so-called "humanized" mouse models) hold great promise, but are often limited by the presence of relatively high levels of endogenous mouse CYPs. Although mouse Cyp genes can be inactivated, one at a time, using a traditional knockout or knock-in approach, the multiplicity of Cyp genes in many of the mouse Cyp subfamilies, and the overlapping substrate specificity of the corresponding CYP enzymes, make the single-Cyp knockout mouse models unsatisfactory as background strains for human CYP-transgenic mice. The specific aim of this Exploratory/Developmental project is to generate a novel mouse strain in which a 1.4-Mb Cyp2a-2b-2f-2g-2s gene cluster of 12 Cyp2 genes and 9 Cyp2 pseudogenes on chromosome 7 is deleted. This mouse strain will provide a clean (or "cleaner") background for crossing with transgenic mouse models expressing one or more of the orthologous human CYP2 genes. The generation of the Cyp2a-2b-2f-2g-2s knockout mouse will pave the way for "humanizing" these important CYP2 genes for a wide variety of in vivo studies, both mechanistic and translational, and will establish a powerful and innovative approach for studying other human CYP as well as non-CYP biotransformation genes, many of which also occur in gene clusters.